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目的探讨miR-15a和miR-16-1寡核苷酸能否增强Raji细胞对阿糖胞苷(cytarabine,Ara-C)的敏感性。方法将化学合成的miR-15a和miR-16-1寡核苷酸利用脂质体2000转染入Raji细胞后,联合Ara-C,CCK8法检测Ara-C的IC50值变化;锥虫蓝细胞计数法检测细胞增殖活性;Hoechst染色观察细胞的凋亡形态;流式细胞仪AnnexinⅤ/PI双染法测凋亡率。结果miR-15a和miR-16-1寡核苷酸转染Raji细胞后,再加入Ara-C,24h Ara-C的IC50值分别为10.41和10.86,明显低于单用Ara-C组(15.43)和随机序列联用Ara-C组(14.92,P<0.05)。锥虫蓝拒染法结果显示,转染后各时间点miR-15a/miR-16-1+Ara-C组较单用miR-15a/miR-16-1组、单用Ara-C组及随机序列+Ara-C组明显抑制了Raji细胞的生长,Hoechst染色可见大量凋亡细胞;流式细胞仪AnnexinⅤ/PI双染色法检测显示,miR-15a+Ara-C组早期凋亡率和晚期凋亡率分别为20.93%和25.27%,miR-16-1+Ara-C组早期凋亡率和晚期凋亡率分别为20.69%和23.13%,均明显高于miR-15a组、miR-16-1组、Ara-C组及随机序列+Ara-C组(P<0.05),后四者的早期凋亡率分别为6.99%、4.73%、10.88%和14.39%,晚期凋亡率分别为10.08%、10.64%、11.83%和11.93%。结论miR-15a和miR-16-1寡核苷酸可增强Raji细胞对Ara-C的敏感性。
Objective To investigate whether miR-15a and miR-16-1 oligonucleotides enhance the sensitivity of Raji cells to cytarabine (Ara-C). Methods The IC50 values of Ara-C were detected by transfection of chemically synthesized miR-15a and miR-16-1 oligonucleotides into Raji cells with Ara-C and CCK8. Cellular proliferation was measured by counting method. Morphological changes of apoptosis were observed by Hoechst staining. Apoptosis rate was determined by flow cytometry Annexin Ⅴ / PI double staining. Results After transfected with miR-15a and miR-16-1 into Raji cells, the IC50 values of Ara-C after 24 h Ara-C treatment were 10.41 and 10.86, respectively, which were significantly lower than those of Ara-C alone group (15.43 ) And random sequence combined Ara-C group (14.92, P <0.05). The results of trypan blue exclusion showed that miR-15a / miR-16-1 + Ara-C at each time point after transfection compared with miR-15a / miR-16-1 alone group and Ara-C alone group The random sequence + Ara-C group significantly inhibited the growth of Raji cells, a large number of apoptotic cells were observed by Hoechst staining. Flow cytometry Annexin Ⅴ / PI double staining showed that the early apoptosis rate and the late apoptosis rate in miR-15a + Ara-C group The apoptosis rates of miR-16-1 + Ara-C group were 20.69% and 23.13%, respectively, which were significantly higher than that of miR-15a group and miR-16 -1 group, Ara-C group and random sequence + Ara-C group (P <0.05). The early apoptosis rates of the latter four groups were 6.99%, 4.73%, 10.88% and 14.39% 10.08%, 10.64%, 11.83% and 11.93% respectively. Conclusion miR-15a and miR-16-1 oligonucleotides enhance the sensitivity of Raji cells to Ara-C.